While the exact mechanisms of ischemia/reperfusion injury remain unclear, our laboratory and others have shown that inhibition of complement activation (e.g., sCR1 or C1 esterase inhibitor), depletion of complement components (e.g., cobra venom factor) and genetically deficient complement component (e.g., C3, C4, MBL, factor D, C1qa) mice have revealed an important role of complement and the specific pathways involved in ischemia/reperfusion injury. During the last funding cycle, we demonstrated that inhibition of mannose binding lectin (MBL) protected the ischemic gastrointestinal system from the primary ischemic event. In contrast, the pulmonary system, which takes a second hit from the primary ischemic gastrointestinal tract, was not protected. Further, complement activation in the lung was observed in C1qa deficient, as well as MBL deficient mice, but not in C2/factor B deficient (KO) mice. Addition of C2 restored the complement activation and tissue inflammation/injury in the C2/fB KO mice following gastrointestinal ischemia/reperfusion (GI/R). These data suggest that a C2 dependent, C1q and MBL independent process is responsible for the complement activation, inflammation and injury following GI/R. Preliminary data, using novel ficolin A KO mice, suggest that this complement activating lectin, ficolin-A, is the initiating molecule responsible for complement activation in the lungs following GI/R. We have also generated novel MBL-A/-C/ficolin-A KO mice for the proposed studies in assessing the role of ficolin-A in pulmonary inflammation and complement deposition following GI/R. Preliminary studies demonstrate biological evidence of a C2 and C4 bypass mechanism for activation of the alternative complement pathway by the MBL complex. One potential molecular mechanism of this bypass involves activation of the coagulation system. Preliminary data demonstrates multiple interactions of the MBL complex with several coagulation proteins. These data demonstrate that simple inhibition of complement in vivo may need a much better understanding of the complex interactions of complement with the coagulation system, an aim of this competitive renewal. In this competitive renewal, we will continue to investigate the molecular mechanisms involved in complement activation during oxidative stress of endothelial cells in vitro and in vivo. The general aim of our laboratory is to characterize the molecular mechanisms governing complement activation during endothelial oxidative stress.